What specific traits of M. tuberculosis (Mtb) are essential for its survival in its infected mammalian hosts? This question can be phrased more specifically: Does Mtb's metabolism change during the course of infection to allow it to persist for long periods during the chronic phase of infections? Previous studies performed in various laboratories including the PI's have shown that many Mtb genes encoding enzymes involved in fatty acid metabolism are upregulated during infections of human macrophages and mice. This information combined with much data in the old and more recent literature suggest a major role for this process in Mtb virulence and that it could provide potential targets for new therapies and diagnostics for TB. This research proposal will study the genetics, biochemistry and physiology of fatty acid metabolism, concentrating on fatty acid degradation and transport, as well as regulators for these and related processes. The rationale for this proposal is that an understanding of the mechanisms of Mtb fatty acid metabolism and how this is important for pathogenesis will be essential for the designing of anti-tubercular strategies that will use components of this process as targets. The aims of the proposal are: I) Specific genes annotated to encode enzymes involved in fatty acid metabolism and their regulators will be identified by a series of approaches and then they will be inactivated;II) The role of these genes and the enzymes/proteins they encode in Mtb virulence will be tested by analyzing the growth and pathogenicity of the mutants in macrophages and mice;III) The function of these genes and the enzymes they encode that are found to be important for M. tuberculosis virulence, as described in specific aim II, will be characterized by purifying the proteins they encode and biochemically assaying the function of the proteins. Other fatty acid metabolic enzymes will also be characterized, even though they may not be essential for pathogenicity as this will provide useful information on Mtb physiology.